Various mass spectrometry techniques are known which attempt to screen for hemoglobin variants.
For example, TW-201211543 (Wu) discloses a quick screening method for a hemoglobin variant comprising: (a) providing a blood sample; (b) quickly separating hemoglobin from the blood sample; (c) determining whether the resulting product of step (b) contains a hemoglobin variant by using Matrix Assisted Laser Desorption Ionization Time of Flight mass spectrometry (MALDI-TOF) in linear mode, and optionally performing the following steps; (d) enzyme hydrolyzing the resulting product of step (b) under ultrasonic vibration; and (e) determining a type of the hemoglobin variant in the resulting product of step (d) by using MALDI-TOF in reflection mode and applying a comparison software. This method further comprises analyzing partial amino acid sequence of the hemoglobin variant by using tandem mass spectrometry (MS/MS) after performing the step (e).
WO 2004/090552 (Dalton) discloses a screening method using mass spectrometry wherein the ionisation technique produces a multiply-charged spectrum and is used to detect variant peptides, polypeptides and proteins that cause or are indicative of disease. The mass spectrometry is focused on a specific multiply-charged species of the polypeptide of interest, enabling a single targeted ionised species and its corresponding variant (if present) to be measured with greater mass precision. An example of use of this method is a screen for sickle cell disease and sickle cell trait by detecting hemoglobin S and other hemoglobin variants using Electrospray mass spectrometry.
However, the known methods are not clinical diagnostic procedures. WO 2004/090552 (Dalton), for example, explicitly states on page 10, line 29 that “this test is not diagnostic.”
Known methods also typically involve complex sample preparation routines. For example, a blood sample may need to be desalted and subjected to liquid chromatography separation which can take a relatively long time to perform.
The article Graça et al., “Electron Transfer Dissociation Mass Spectrometry of Hemoglobin on Clinical Samples”, J. Am. Soc. Mass Spectrom. (2012) 23:1750-1756, discloses a method for identifying hemoglobin variants from whole blood using Electron Transfer Dissociation (“ETD”) mass spectrometry.
The article Acosta-Martin et al., “Quantitaive Mass Spectrometry Analysis of Intact Hemoglobin A2 by Precursor Ion Isolation and Detection”, Anal. Chem. (2013), 85:7971-7975, discloses a method of quantifying hemoglobin using ETD mass spectrometry.
The article Edwards et al., “Top-Down Proteomics and Direct Surface Sampling of Noenatal Dried Blood Spots: Diagnosis of Unknown Hemoglobin Variants”, J. Am. Soc. Mass Spectrom. (2012), 23:1921-1930, discloses a method for screening for hemoglobin variants using ETD mass spectrometry.
The techniques described in these articles work with denatured (c.f. native) hemoglobin and therefore require complex sample preparation routines.
Fast methods of obtaining a clinical diagnosis of native hemoglobin variants using mass spectrometry are not known.
It is desired to be able to provide an improved method of screening or testing for native hemoglobin variants using mass spectrometry.